The mammary gland is a complex organ whose growth and development are controlled by the interaction of a wide variety of hormones and growth factors also involved in the etiology and progression of the cancerous state. Our emphasis has been on the interactions of prolactin (Prl), estrogens (E), and progesterone (P), with recent work also examining how epidermal growth factor (EGF), and transforming growth factors alpha and beta are affected by the interplay of these three classical hormones. Lobulo-alveolar development of the mammary gland requires the priming action of both E and P to induce EGF receptors and production of EGF- like growth factors. E induces the P receptor at branch points of the ductal tree; P induces the Hox genes and promotes branching morphogenesis. In concert with insulin, Prl and glucocorticoids, EGF or TGF-alpha can promote full lobulo-alveolar development in vitro. Programmed cell death occurs within 3 days of removal of Prl. The apoptosis of alveolar cells is regulated, in part, by p53. Prl induced growth of NOG-8 normal mouse mammary epithelial and T47D human breast cancer cells involves activation of ras, raf-1, MEK and MAP kinases within 1 to 5 min. JAK2 and SHC are phosphorylated within 1 min. and associated with Grb2 and Sos. Stat 5 is phosphorylated within 30 min. of Prl treatment. Prl induced growth of human breast cancer cells can be blocked by non-steroidal antiestrogens such as tamoxifen (TAM) and the steroidal pure antiestrogens ICI182780 and ICI164384. This action occurs directly through the Prl receptor. This was confirmed by transfection of the genes for either the long or the short form of the Prl receptor into NIH 3T3 cells. Both forms of the receptor transmit the growth signal while only the long form promotes differentiation. Transfectants carrying either form of the receptor acquire the ability to bind TAM as well as Prl and the Prl binding is blocked by the antiestrogens. Neonatal exposure to TAM results in an adult mouse with poorly developed mammary glands containing more pre-neoplastic hyperplastic alveolar nodules. Serum Prl levels in the treated animals are constitutively elevated . The antiprolactin action of TAM may have important clinical implications for ER-/PrlR+ tumors. The importance of Prl in human breast disease was confirmed by studies in MCF-7 and T47Dco cells which show that biologically active Prl is synthesized and secreted by these human breast cancer cells. Growth of both ER- and ER+ human breast cancer cells is inhibited by 70-85% by anti-Prl antibodies. Conditioned media and extracts from cells precipitated with anti-Prl antibodies, contain a single band of approximately 22kDa, the size of pituitary Prl. The breast cancer cells synthesize and secrete 0.7ug/ml of biologically active Prl which is about 30% of the level produced by pituitary cells similarly cultured. By RT-PCR, 80% of human breast cancer cell lines and 11 of 12 breast cancer biopsies have Prl message. Cancerous tissue has significantly more Prl and Prl receptor message than adjacent, non-involved tissue from the same patient. These data suggest an autocrine role of Prl in human breast cancer which may have implications in therapy.